Edwin A. Dawes

Accumulation of a poly(hydroxyalkanoate) copolymer containing primarily 3-hydroxyvalerate from simple carbohydrate substrates by Rhodococcus sp. NCIMB 40126☆

A number of taxonomically-related bacteria have been identified which accumulate poly(hydroxyalkanoate) (PHA) copolymers containing primarily 3-hydroxyvalerate (3HV) monomer units from a range of unrelated single carbon sources. One of these, Rhodococcus sp. NCIMB 40126, was further investigated and shown to produce a copolymer containing 75 mol% 3HV and 25 mol% 3-hydroxybutyrate (3HB) from glucose as sole carbon source. Polyesters containing both 3HV and 3HB monomer units, together with 4-hydroxybutyrate (4HB), 5-hydroxyvalerate (5HV) or 3-hydroxyhexanoate (3HHx), were also produced by this organism from certain accumulation substrates. With valeric acid as substrate, almost pure (99 mol% 3HV) poly(3-hydroxyvalerate) was produced. N.m.r. analysis confirmed the composition of these polyesters. The thermal properties and molecular weight of the copolymer produced from glucose were comparable to those of PHB produced by Alcaligenes eutrophus.

A survey of the accumulation of novel polyhydroxyalkanoates by bacteria

SummaryA wide range of bacterial strains were examined for their ability to accumulate polyhydroxyalkanoates (PHA) from various carbon sources. Strains were selected from those reported to accumulate poly-3-hydroxybutyrate (PHB), related organisms and laboratory stocks. Other strains known to utilize n-alkanes, n-alcohols or n-acids were chosen to investigate their ability to produce long-chain PHAs. Five strains accumulated only PHB, 13 accumulated PHAs containing only C4 and C5 units and 7 accumulated PHAs containing 3-hydroxyacid units in the range C5 to C10.

Biosynthesis and composition of bacterial poly(hydroxyalkanoates)

It is well established that Alcaligenes eutrophus can accumulate a copolymer containing 3-hydroxybutyrate and 3-hydroxyvalerate, but longer 3-hydroxyacid monomers have not been reported to occur in this organism. The properties of the enzymes of poly(hydroxyalkanoate) (PHA) biosynthesis are discussed and it is proposed that the substrate specificity of the polymerizing enzyme restricts the range of monomer units incorporated into PHA. Various other bacteria produce similar copolymers from propionic acid and/or valeric acid. A number of Pseudomonas species accumulate PHAs containing longer-chain monomer units from linear alkanoic acids, alkanes and alcohols.

The role of glucose limitation in the regulation of the transport of glucose, gluconate and 2-oxogluconate, and of glucose metabolism in Pseudomonas aeruginosa.

The pathway of glucose metabolism in Pseudomonas aeruginosa was regulated by the availability of glucose and related compounds. On changing from an ammonium limitation to a glucose limitation, the organism responded by adjusting its metabolism substantially from the extracellular direct oxidative pathway to the intracellular phosphorylative route. This change was achieved by repression of the transport systems for gluconate and 2-oxogluconate and of the associated enzymes for 2-oxogluconate metabolism and gluconate kinase, while increasing the levels of glucose transport, hexokinase and glucose 6-phosphate dehydrogenase. The role of gluconate, produced by the action of glucose dehydrogenase, as a major inhibitory factor for glucose transport, and the possible significance of these regulatory mechanisms to the organism in its natural environment, are discussed.

Effect of carbon source and concentration on the molecular mass of poly(3-hydroxybutyrate) produced by Methylobacterium extorquens and Alcaligenes eutrophus

In shake-flask culture, Methylobacterium extorquens accumulated poly(3-hydroxybutyrate) (PHB) possessing a substantially higher weight-average molecular mass (MW) than previously reported for this organism. The MW of PHB produced by M. extorquens was dependent on the initial concentration of methanol or sodium succinate, used as sole carbon sources. The highest MW values (0.6 and 1.7 × 106) were obtained with low initial concentrations of methanol or sodium succinate (4.0 and 3.0 g l−1, respectively) and the latter substrate always yielded PHB of higher MW than that produced from methanol. Thus PHB with an MW in the range 0.2–1.7 × 106 could be produced by selection of the carbon source and its concentration. In contrast to the findings with M. extorquens, the MW of PHB produced by Alcaligenes eutrophus was high (1.1–1.6 × 106) and generally unaffected by the choice or concentration of the carbon source. The use of glycerol as sole carbon source did, however, result in the accumulation of PHB with a markedly lower MW (5.5–8.5 × 105) than that produced from other sole carbon sources by this organism under similar conditions.

Polyhydroxybutyrate: an intriguing biopolymer.

The microbial polymer poly-3-hydroxybutyrate (PHB) and related poly-hydroxyalkanoates, such as poly-3-hydroxyvalerate and poly-3-hydroxyoctanoate, are unique biodegradable thermoplastics of considerable commercial importance. The structure, properties and regulation of synthesis and degradation of PHB are reviewed and the microbial production of copolymers of 3-hydroxybutyrate and 3-hydroxyvalerate, with properties varying according to copolymer composition, is discussed.

Effects of Carbon Source and Inorganic Phosphate Concentration on the Production of Alginic Acid by a Mutant of Azotobacter vinelandii and on the Enzymes Involved in its Biosynthesis

The specific activities of the key enzymes involved in the biosynthesis of the exopolysaccharide alginic acid by Azotobacter vinelandii were determined in extracts of batchcultured organisms grown with different carbon sources in the presence of limited and excess inorganic phosphate. Alginic acid production was also measured. Glucose, fructose, sorbitol, mannitol, glycerol and gluconate resembled sucrose in supporting much greater alginate production in media containing growth-limiting amounts of inorganic phosphate. Mannose supported only poor growth with no alginate formation, and growth did not occur on acetate. Increases in the specific activities of phosphomannose isomerase, GDPmannose pyrophosphorylase and GDPmannose dehydrogenase were accompanied by increased alginic acid production. Our results accord with the suggestion that alginate formation is controlled by derepression of key biosynthetic enzymes.

Growth and Survival of Bacteria

The survival of a bacterium in its natural habitat depends on its ability to grow at a rate sufficient to balance death caused by starvation and other natural causes such as temperature, pH, and osmotic fluctuations, as well as predation and parasitism. In discussing survival under extreme conditions, Shilo (1979) has drawn attention to the difference between (1) stable ecosystems (exemplified by the continuous high temperatures in thermal springs, continuous high salinity as in the Dead Sea, and continuous high hydrostatic pressure typical of the ocean depths), which are inhabited by organisms with narrow adaptations; and (2) fluctuating ecosystems (typified by marshes, swamps, and shallow lakes with pronounced diurnal fluctuations of physical and chemical parameters) that harbor organisms with a much greater versatility of response.

The Production of Polyhydroxyalkanoates from Unrelated Carbon Sources

Several bacteria have been found to accumulate polyhydroxyalkanoates (PHAs) containing 3-hydroxyvalerate (3HV) and 3-hydroxybutyrate (3HB) monomers from glucose and other carbon sources. These bacteria are members of the taxonomically related genera Rhodococcus, Nocardia and Corynebacterium. The proportion of 3HV and 3HB monomers present in PHA is dependent on the carbon source but 3HV is generally the major 3-hydroxyacid.

Studies on Some Enzymes of Alginic Acid Biosynthesis in Azotobacter vinelandii Grown in Continuous Culture

Summary: When a mutant of Azotobacter vinelandii was grown in continuous culture the amount of exocellular polysaccharide produced was dependent on both the dissolved oxygen tension (d.o.t.) and the carbon source: sucrose supported alginate synthesis in phosphate-limited medium whereas sorbitol did not. Changes in the specific activities of two of the key enzymes of alginate biosynthesis (phosphomannose isomerase and GDPmannose pyrophosphorylase), measured in extracts of cells grown with sucrose under a range of d.o.t. values, were reflected by the observed changes in alginate production; the activity of GDPmannose dehydrogenase was unchanged. A similar correlation between the specific activities of these enzymes and the rate of alginate production was observed during a transition from sorbitol to sucrose as the sole carbon ource, but in this experiment the activity of GDPmannose dehydrogenase also increased with increasing alginate production. After prolonged continuous cultivation on sucrose the mutant gradually lost the ability to produce alginate. The key enzymes of alginate biosynthesis could not be detected in extracts of this non-alginate-producing strain, which had also lost the ability to encyst. These results support the suggestions that alginate formation is controlled by derepression of key biosynthetic enzymes and that alginate plays an important role in encystment.